WO 2003/048300 discloses a method for treating cells when a flow cytometer or the like is applied for detection in a fluorescence in situ hybridization method (FISH method). In the FISH method, first, a pretreatment for hybridizing a fluorescently labeled probe to the base sequence of a target site present in the nucleus of a cell is performed to fluorescently label the target site. Subsequently, a fluorescence signal (bright spot) generated from the fluorescently labeled probe is detected.
In the FISH method, the fluorescence image is captured with a fluorescence microscope, an imaging flow cytometer, or the like. However, even in the case where the same fluorescently labeled probe is used, the brightness may be different between bright spots in one fluorescence image due to variations in pretreatment. Also, since cells take a three-dimensional shape in a flow cell of a flow cytometer, the brightness of a plurality of bright spots in one captured fluorescence image may be different from each other depending on the position (depth) of the bright spot when light is radiated thereto, for example, depending on whether the bright spot is present on the surface of the nucleus or near the center of the nucleus. As described above, the brightness of bright spots may vary in one fluorescence image when a fluorescence image of a cell is captured in a flow cytometer. Therefore, when an operator checks the presence or absence of a bright spot in the fluorescence image captured by a flow cytometer in the FISH method, there is a case where a darker bright spot cannot be detected.
Furthermore, in a multicolor FISH method, by using a probe labeled with a green fluorescent dye and a probe labeled with a red fluorescent dye, the presence or absence of a fused bright spot in which the two probes are combined by being adjacent to each other and in which yellow fluorescence is emitted is detected. Specifically, a fused bright spot is detected by superimposing a fluorescence image obtained by imaging the green fluorescent dye and a fluorescence image obtained by imaging the red fluorescent dye. However, as described above, in one fluorescence image captured in a flow cytometer, the brightness of a plurality of bright spots may vary. Therefore, there is a problem that, when confirming a fused bright spot in a fluorescence image captured by using a flow cytometer in the multicolor FISH method, for example, in the case where a certain red bright spot is dark and a green bright spot adjacent thereto is bright, the red bright spot is overwhelmed by the green bright spot and the operator cannot determine that the bright spot is a fused bright spot when two fluorescence images are superimposed.